The global health burden ofthe major hemoglobin disorders, sickle cell anemia and B-thalassemia, is enormous and predicted to grow. Increased fetal hemoglobin (HbF) greatly ameliorates the morbidity and mortality of these disorders. A long sought goal is directed reactivation of HbF expression in adults with hemoglobin disorders. Largely through human genetic studies, substantial progress has been made in understanding the regulatory factors controlling the fetal (a2Y2) to adult (a2B2) switch and how HbF silencing is maintained in the adult. A premise of the proposed work is that fundamental findings on hemoglobin switching provide a platform for establishing mechanism-based approaches to the identification of small molecules that induce HbF in adult erythroid cells. If successful in this overall goal, new therapies may be developed for treatment of patients with the major hemoglobin disorders. This proposal encompasses three independent, but interrelated, aims. First, the mechanisms by which the gene encoding BCL11A, a central repressor of HbF expression, is regulated will be explored using high-resolution nascent transcription mapping, chromatin occupancy, and assays of putative cis-regulatory elements. Down-regulation of BCL11A expression or function provides a direct route to relief from HbF silencing. An additional goal of this aim isalso to determine how genetic variation in the BCL1 IA gene influences expression of BCL11A itself, as this knowledge may lead to approaches to direct down-regulation of BCL11A expression. Second, high throughput screening for small molecule inducers of HbF will be performed in cultured erythroid cells, and promising hits will be evaluated further for the pathways affected and for optimization as therapeutics. Third, screens with genome-wide short hairpin RNA (shRNA) and open reading frame (ORF) libraries will be performed to identify novel genes/pathways for induction of silenced HbF in primary human CD34 derived erythroid progenitors. Through the multidisciplinary approaches in this proposal basic findings on HbF regulation will be translated to advance identification and development of new therapeutics for patients with hemoglobin disorders.